Exogenous secretory peptides expressed in salivary glands are of potential clinical importance in the regulation of salivary flow and composition, or as therapeutic peptides in the mouth and gastrointestinal tract. Adenovirus-mediated gene transfer to salivary glands has recently been demonstrated, suggesting that the expression of therapeutic peptides in salivary glands and saliva is clinically feasible. Controlling how therapeutic peptides are released is important for the clinical success of this approach. Ideally, the release of such peptides can be regulated according to their therapeutic role so that they are released either by constitutive secretion or by regulated secretion of peptides stored in secretory granules. The signals required for targeting or sorting of peptides to the regulated secretory pathway in exocrine cells are not known. The long term goal of the present research is to determine the amino acid sequence(s) and mechanisms required for targeting of peptide hormones to the regulated secretory pathway in exocrine cells. Exocrine and endocrine cells express both a regulated and a constitutive secretory pathway. The regulated pathway is distinguished from the constitutive pathway, common to all eukaryotic cells, by slower release times (in the absence of stimulation), intracellular storage of secretory proteins and stimulated secretion in response to secretogogues. Regulated secretory proteins, including digestive enzymes and peptide hormones are targeted or sorted to the regulated pathway in the trans-Golgi network or maturing secretory granules. It appears that a common sorting mechanism exists for different endocrine cells since foreign endocrine proteins are correctly sorted in transfected endocrine cells. The sorting mechanisms that function in exocrine cells are not known. The peptide hormone atrial natriuretic factor (ANF) is expressed in salivary gland acinar cells and, thus, represents a physiologically relevant model for these studies. In the present proposal we will test the hypothesis that proANF is sorted to the regulated pathway in exocrine cells and then elucidate the sorting signal required. The following specific aims are proposed: 1) Determine if salivary gland-derived cell lines exhibit regulated secretion. 2) Determine if proANF is sorted to the regulated secretory pathway in exocrine cells. 3) Identify the amino acid sequences of proANF that play a role in sorting of the protein to the regulated secretory pathway. 4) Determine the sorting mechanism for proANF: Determine if the sorting signals are required for aggregation of proANF or interaction with a cellular sorting receptor.